Project Summary/Abstract The K22 applicant, Dr. Steven Sizemore, obtained his Ph.D. in the laboratory of Dr. Graham Casey at the Cleveland Clinic Foundation. His dissertation research focused on the fundamental mechanisms underlying breast cancer metastasis. The candidate's training continued as a Department of Defense CDMRP Breast Cancer Research Program supported postdoctoral fellow at Case Western Reserve University. This work concentrated on identifying the molecular drivers of basal-like breast cancer. Currently, the candidate is a Senior Research Associate at The Ohio State University James Comprehensive Cancer Center (OSUCCC). The OSUCCC is ranked by the National Cancer Institute as an ?exceptional? center and provides a vibrant and collaborative environment in which the candidate has continued his training towards his long-term goal of becoming tenure-track faculty at a Research I level university. The candidate's current work, on which this proposal is based, centers on identifying the master regulators of soft tissue sarcoma (STS) metastasis and is supported through a seed grant from the OSUCCC. STS is a diverse collection of cancers of mesenchymal origin. The 5-year survival for patients with metastatic STS is a dismal 16% and there are few treatment options for these patients. The long-term objective of this research is to address the urgent, unmet need for a better understanding of the key molecular pathways that drive metastasis in STS and identify inhibitors of these pathways to improve the treatment options for advanced STS. In pursuit of this objective, the candidate identified the phosphatase subunit PPP2R1B as a suppressor of STS metastasis and one of its phosphoprotein targets, RALA, as a putative actionable target for treating advanced STS. This proposal will thoroughly test the hypothesis that RALA, and the closely related RALB, are important and targetable drivers of STS tumor growth and metastasis that regulate STS progression, at least in part, by controlling exosome production. Specific Aim 1: will test the functional requirement of the RAL isoforms and their effectors in STS growth and metastasis in vitro and in vivo by coupling genetic silencing of these isoforms with rescue experiments utilizing specific interaction-deficient RAL mutants. Specific Aim 2: will investigate the essential requirement of the RAL isoforms and their effectors in exosome production by STS cells and determine if these RAL-dependent exosomes control tumor growth and metastasis. Specific Aim 3: will utilize STS cell lines and patient derived xenografts to determine the effectiveness of RAL-targeting inhibitors as innovative treatments for STS. Successful completion of this proposal will identify an important actionable target for improved treatment of advanced STS and identify a novel mechanism regulating cancer exosomes. This award will allow the candidate the protected time required to transition his research from a breast cancer- focus to the understudied field of STS metastasis and acquire the additional training required to become a leader in the field of cancer-associated exosomes.